Project Summary The discovery of antibiotics to treat bacterial infections has had a dramatic and positive impact on human health. However shortly after the introduction of new antibiotics, resistance often develops. The emergence and spread of antibiotic resistant bacteria is considered a worldwide health emergency. In addition to a public health threat, antibiotic resistant bacteria also place a significant financial burden on the health care system. It is clear new methods to target antibiotic resistance are seriously needed. The bacterial cell envelope is essential for cell viability and is the target of many of the most commonly used antibiotics including ?-lactam antibiotics like penicillin. These are general broad-spectrum antibiotics that target peptidoglycan biosynthesis by inhibiting the transpeptidase activity of penicillin binding proteins. Our long-term goal is to understand the mechanism by which ?-lactams are sensed by Gram- positive bacteria. Organisms in the Bacillus cereus group including Bacillus thuringiensis and Bacillus anthracis are highly resistant to ?-lactam antibiotics. This resistance is dependent upon the Extra- Cytoplasmic Function (ECF) ? factor ?P, which we have found is specifically activated by a subset of ?- lactam antibiotics. ECF ? factors represent an important class of signal transduction systems which compared to other regulatory systems are relatively poorly understood. We have found that ?P is activated upon the sequential proteolytic destruction of the anti-? factor RsiP. Our data indicate that BT3488 a penicillin binding protein is required for site-1 cleavage of RsiP. These findings raise several important questions: How are ?-lactam antibiotics sensed by the cell? What factors are required for activating ?P in response to ?-lactam antibiotics? Here we propose to 1) Determine the role of BT3488 and RsiP in sensing ?-lactams and controlling ?P activation and 2) Identify additional factors required for ?P activation.